Fig. 4: Apical polarization in PrE cells is required for directed migration and sorting.
From: Coupling of cell shape, matrix and tissue dynamics ensures embryonic patterning robustness
a, Immunofluorescence image of a 3× blastocyst at stage E3.75 showing laminin distribution around PrE cells. White dotted line denotes the ICM–cavity interface; white arrowhead indicates a GATA6-expressing PrE cell. b, Immunofluorescence image of a 3× blastocyst at stage E3.75 showing PKCλ+ζ distribution in PrE cells. White arrowhead denotes the leading edge of a PrE cell with PKCλ+ζ localization. c, Immunofluorescence image of an E3.75 ICM showing PKCλ+ζ localization in PrE and EPI cells. White dotted lines denote cell boundaries; the yellow line marks the segment from the cell inner edge (towards ICM centroid) to the cell outer edge (towards the ICM–fluid interface) along which fluorescence intensity is measured. d, Normalized fluorescence intensity of PKCλ+ζ in individual inside cells from E3.75 isolated ICMs. n = 260 cells from 32 ICMs. Each of the thin lines corresponds to measurement from one cell. Bold line and shaded region indicate mean ± s.d. of aPKC intensity for GATA6-high and GATA6-low cells. e, Left, schematic of polarization index. Right, boxplots for comparison of the polarization index in PrE (GATA6-high) versus EPI cells (GATA6-low). GATA6 expression level is categorized as high or low by thresholding the bimodal distribution of GATA6 fluorescence intensity. n = 136 GATA6-high and 124 GATA6-low cells from 32 ICMs. One-way ANOVA, P = 6.03 × 10−20. f, Scatter-plot of polarization index of cells versus radial distance of the cell from the ICM centroid. Black dotted line indicates linear regression with Pearson’s R = 0.079, P = 0.205. n = 260 cells from 32 ICMs. g, Immunofluorescence images of control and Gö6983-treated E4.0 isolated ICMs (left) and quantification of sorting score (right). n = 16 and 24 ICMs for control and Gö6983-treated ICMs, respectively. Two-sided independent-samples t-test, P = 8.01 × 10−4. h, Immunofluorescence images of representative WT, Prkci+/+Prkcz−/− and Prkci+/−Prkcz−/− E4.5 blastocysts (left) and quantification of number of ectopic PrE cells in E4.5 blastocysts from each group (right). n = 25, 17 and 14 blastocysts for WT, Prkci+/+Prkcz−/− and Prkci+/−Prkcz−/−, respectively. Two-sided Mann–Whitney U-test, P = 2.43 × 10−4 and 6.36 × 10−4. Scale bars, 20 μm. *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001.
